Anti gbp1

Antibodies were purchased from the following sources: anti-CD99 FITC conjugated (BD Biosciences, catalog no: 555688, concentration 1:20), anti-FLI1 (Abcam, catalog no: 133485, concentration 1:2,000), anti-BARD1 (Bethyl Laboratories, catalog no: A300–263A, 1:2,000), anti-BARD1 (Abcam, catalog no: ab50984, concentration 1:100), anti-GBP1 (Abcam, catalog no: 131255, concentration 1:300 for IHC), anti-GBP1 (Santa Cruz Biotechnology, catalog no: sc-53857, concentration 1:200 for Western blot analysis), anti-phospho (Ser 139)-γH2A.X (Millipore Sigma, catalog no: 05–636, concentration 1:2,500 for immunofluorescence), anti-phospho (Ser 139)-γH2A.X (Invitrogen, catalog no: MA1–2022, concentration 1:1,000 for Western blot analysis), goat anti-mouse IgG AF-488 (Thermo Fisher Scientific, catalog no: A-11001, concentration 1:2,000), tubulin (Cell Signaling Technology, catalog no: 2144S, concentration 1:5,000), vinculin (Cell Signaling Technology, clone E1E9V, catalog no: 13901S, concentration 1:5,000), and anti-rabbit IgG-horseradish peroxidase (HRP) (Promega, catalog no: W401B). Additional specialized reagents include: BMN 673 (talazoparib; Cayman Chemical, catalog no: 19782), MK-4827 tosylate (niraparib; Cayman Chemical, catalog no: 20842), DMSO (MP Biomedicals, catalog no: 196055), doxorubicin (Cayman Chemical, catalog no: 15007), and IFNγ (R&D systems, cat no: 285IF100).

BFP reporter gene expression was monitored to determine HIV-1 LTR and LTR12C activity in transfected HEK293T cells or electroporated CD4⁺ T cells. Two days post transfection or electroporation, cells were washed in PBS with 2% FCS, fixed in 4% PFA and fluorescence was determined using a BD FACS Canto II flow cytometer. To quantify GBP protein levels, cells were permeabilized using the FIX & PERM kit (Nordic-MUbio, #GAS-002–1) according to the manufacturer's instructions. Subsequently, cells were stained using anti-GBP1 (Santa Cruz, #sc-53857), anti-GBP2 (Origene, #TA500657), anti-GBP5 (Santa Cruz, #sc-160353) and the following secondary antibodies: goat anti-rat, APC-conjugated (life technologies, #A10540), goat anti-mouse, PE-conjugated (life technologies, #P852), donkey anti-goat, Alexa Fluor 647-conjugated (life technologies, #A21447)

Cell and tissue samples were lysed using RIPA buffer (Sigma-Aldrich, Saint Louis, MO) supplemented with protease inhibitor mixture (Roche Diagnostics). Protein concentrations were measured using the bicinchoninic acid (BCA) method. Equal amounts of proteins were loaded and separated on 10% SDS-PAGE, transferred to a nitrocellulose membrane and probed with indicated antibodies. Blots were visualized by enhanced chemiluminescence (Thermo Scientific, Rockford, IL). The following antibodies were used in this study: anti-Gbp1, anti- NFκB and anti-Tom20 (Santa Cruz); anti-phospho-NFκB, anti-phospho-p53, anti-p53, anti-phospho-AMPK, anti-AMPK, anti-Parkin, anti-p62, anti-LC3I/II, anti- β-actin and anti-tubulin (Cell Signaling Technology); anti-Lcn2 (R&D System, Minneapolis, MN).